1. Field of the Invention
The present invention relates to recombinant plasmids and to bacteria containing the recombinant plasmids which produce polysaccharide depolymerase, wherein the recombinant plasmid contains a DNA segment of a bacteriophage for Erwinia amylovora. In particular the present invention relates to Escherichia coli containing a recombinant plasmid with a DNA segment of phage ERA103 which expresses a depolymerase.
2. Prior Art
Erwinia amylovora is recognized as the causitive agent of fireblight in rosaceous plants (Ayers, A. R., et al., Appl. Environ. Microbiol. 38: 659-666 (1979)). The pathogenic strains of this bacteria have been demonstrated to produce a polysaccharide capsule which has been implicated in xylem vessel occlusion and plasmolysis of xylem parenchymal cells, which are symptomatic of the disease. U.S. patent application Ser. No. 662,056, filed Oct. 18, 1984 now U.S. Pat. No. 4,678,750 describes the use of a bacteriophage induced depolymerase from Erwinia amylovora for the treatment of this disease.
Bacteriophage encoded polysaccharide depolymerases have been described for many bacterial genera (Adams, M. H., et al., Virology 2: 719-736 (1956) and Higashi, S., et al., J. Gen. Appl. Microbiol. 24: 143-153 (1978)). Bacteriophage PEal (h) has been shown to degrade to the polysaccharide capsule of Erwinia amylovora (Hartung, J. S., et al., Phytopathology 72: 945 (1982)). Bacteriophage ERA103 has been found to infect the plant pathogen E. amylovora NCPPB595 and produce a depolymerase that degrades the polysaccharide capsule of this bacteria. The problem is that it is difficult and expensive to isolate significant amounts of the depolymerase by induction with the bacteriophage.
Much more of the depolymerase might be obtained if it was possible to clone a segment of the phage DNA encoding for the depolymerase into a vector plasmid. It was uncertain whether the depolymerase could be produced without the presence of the host Erwinia amylovora or whether the genes responsible for encoding the depolymerase could be cloned into a recombinant plasmid.
Objects
It is therefore an object of the present invention to provide a recombinant plasmid in a bacteria which encodes for the expression of a depolymerase. Further the present invention relates to a recombinant plasmid including a segment of a bacteriophage which infects Erwinia amylovora and which encodes for the depolymerase. In particular it is an object of the present invention to provide a recombinant plasmid including a segment of bacteriophage ERA 103 which encodes for the expression of the depolymerase in E. coli. Finally, it is an object of the present invention to provide a method for producing the depolymerase using the recombinant plasmid which is simple and economical. These and other objects will become increasingly apparent by reference to the following description and the drawings.